Epithelial‐to‐Mesenchymal Transition Induced by Accumulation of Autophagosomes in Podocytes

Abstract

Mature podocytes are highly differentiated cells in the kidney and changes in this differentiated status of podocytes such as epithelial‐to‐mesenchymal transition (EMT) or dedifferentiation contribute to the development of glomerular injury or sclerosis. The present study was designed to test the hypothesis that EMT occurs in podocytes if its autophagic process is impaired. In conditionally immortalized mouse podocytes, we demonstrated that blockade of autophagic flux by lysosomal V‐ATPase inhibitor, bafilomycin A1 or by silencing of its isoform using V‐ATPase V0 siRNA significantly induced EMT, as shown by decreased expression of epithelial markers, p‐cadherin and ZO‐1, and increased expression of mesenchymal markers, FSP‐1 and α‐SMA. This enhanced EMT was accompanied by reduction of podocin production, a podocyte function. All these changes were confirmed by confocal microscopy, real time RT‐PCR and Western blot analysis. Correspondingly, both bafilomycin A1 and V‐ATPase V0 siRNA blocked autophagic flux that resulted in a large increase in autophagosomes in podocytes, as shown by increased expression of microtubular associated protein 1 light chain 3‐II (LC3B‐II). In particular, a dramatic increase in the level of cytoplasmic scaffold protein P62 was observed when autophagic flux was blocked. In contrast, a specific inhibitor of autophagosome formation, spautin‐1 had no effect on podocyte EMT under control condition, but it significantly reversed bafilomycin A1‐induced effects on both EMT and LC3B‐II or P62 expression. These results suggest that the maturation and normal function of podocytes are critically controlled by autophagic process and that lysosome dysfunction and consequent impairment of autophagic flux may induce EMT in these podocytes (supported by NIH grants HL057244, HL‐75316 and DK54927).